This is an application for an R21 grant to explore a novel approach to the in vivo analysis of antigen receptor function. Antigen receptor transgenic mice have been extraordinarily useful in the analysis of immune specificity, but they also pose a series of limitations to certain analyses. Taking advantage of these antigen receptor transgenic systems to screen the effects of various knockout mutations has been time-consuming and cumbersome, primarily because of the need for introducing the transgenes onto the knockout genetic background. In this proposal we suggest a novel general strategy that we will apply to two particular questions in B lymphocyte biology. The methods are, however, applicable to the analysis of several lymphocyte subsets and to the study of a wide range of receptor-mediated processes. The strategy involves the development of single- chain and double-chain antibodies with specificity for constant regions or framework regions of antigen receptor proteins. The genes encoding these proteins will be used to generate transgenic mice that express these proteins as membrane antigens on the surface of selected tissue cells. The feasibility of using these mice to analyze receptor-mediated processes will then be tested in vivo. The long-term goal of this work is to generate a general approach to exploit the mutational analysis of receptor-signaling. These studies should have broad applications to questions of lymphocyte activation, differentiation, and survival.